A Sayadi1, J Jeyakani2, S H Seet1, C-L Wei2, G Bourque2, F A Bard1, N A Jenkins1,3, N G Copeland1,3and E A Bard-Chapeau1,4
1 Institute of Molecular and Cell Biology, Singapore, Singapore
2 Genome Institute of Singapore, Singapore, Singapore
3 Current address: The Methodist Hospital Research Institute, Houston, TX, USA.
4 Current address: Novartis Institutes for Biomedical Research, WKL-125.1.05A, Basel CH-4002, Switzerland.
Oncogene advance online publication 3 August 2015; doi: 10.1038/onc.2015.286
MECOM gene encodes several transcription factors variants including MDS1-EVI1, EVI1, and EVI1Δ324. While MDS1-EVI1 has been associated with tumor suppressing activity, EVI1 is a known oncogene in various cancers, whose expression is associated with poor patient survival. Although EVI1Δ324 is co-transcribed with EVI1, its activity in cancer cells was not fully understood. Previous reports described that, unlike EVI1, EVI1Δ324 protein cannot transform fibroblasts, because of its disrupted N-terminal ZNF domain.
To better understand EVI1Δ324 biology and function, we obtained another genome-wide binding occupancies and expression data in ovarian cancer cells and compared it to the one we obtained previously for EVI1 (Proc Natl Acad Sci U S A. 2012;109(6):2168-73). We characterized EVI1Δ324 DNA binding sites, binding motif and target genes. Comparative analyses show that EVI1 and EVI1Δ324 share similar transcriptional activities linked to their common C-terminus ZNF domain. They bind to an ETS-like motif, target to a large extent the same genes, and cooperate with AP1 transcription factor. 70.7% of EVI1Δ324 occupied genes were also found as EVI1 bound genes. More strikingly, EVI1 and EVI1Δ324 differentially expressed genes were 99.87% identical, indicating comparable transcriptional regulatory functions. Consistently with gene ontologies linked to these target genes, EVI1Δ324 expression in HeLa cells could enhance anchorage-independent growth, like EVI1, showing that EVI1Δ324 expression also lead to pro-oncogenic effects.
The main specific feature of EVI1 variant is its N-terminus ZNF domain that binds DNA through GATA-like motif. We found that most GATA-like EVI1 ChIP-Seq peaks are far from genes and not involved in transcriptional regulation. These genomic regions were enriched in simple sequence repeats and displayed high meiotic recombination rates. Thus, the suspected transforming activity of the EVI1 N-terminus ZNF domain could be linked to a role in the regulation of homologous recombination rather than specific transcriptional changes. In line with this, our previous mass spectrometry analysis (Proc Natl Acad Sci U S A. 2013;110(31):E2885-94) reported interactions of EVI1 with proteins involved in DNA damage repair, DNA recombination, and meiosis in SKOV3 and Hela cells. Overall, these findings hint a direct involvement of EVI1 in regulating genomic instability through the protein recruitment at specific genomic sites.
Figure legend: Genomic features of EVI1 and EVI1Δ324 isoforms
EVI1 contains two separated DNA binding zinc finger domains. The first domain is localised at the N-terminal part and recognize a GATA-like motif, the second domain is at the C-terminal part and recognize an ETS-like motif. EVI1Δ324 is a shorter isoform, characterized by the disruption of the N-Terminal ZNF domain. The loss of two zing finger motifs makes EVI1Δ324 N-terminal ZNF domain inactive for DNA binding. On the other hand, EVI1Δ324 C-Terminal ZNF domain is kept active and able to occupy an ETS-like DNA binding motif. In this study, we observed that EVI1 and EVI1Δ324 display target genes in common, and their binding sites are often located near an AP1 motif. EVI1 N-terminal domain, on the other hand bound genes far from genes and in regions associated with high recombination rate.